Protein modification by 4-hydroxynonenal (HNE) modulates cell function: identification of specific proteins as cellular targets of HNE binding

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This research program is aimed at clarifying the role of oxidative stress in proteins modification in isolated cells. Recently, hydroxynonenal (HNE) have emerged as the best available markers of oxidative stress-induced lipid peroxidation (LPO) in vivo. HNE is an aldehyde end-product generated by peroxidation of the most abundant class of n-6 polyunsaturated fatty acids. Similar to free radicals, HNE is electrophiles that react readily to proteins to form HNE-protein adducts, but its relatively longer half-life makes it candidates for the propagation of the damage to neighboring cells. The interest for HNE stems not only from its potential use as a biomarker of oxidative stress-induced LPO, but also because of accumulating evidence indicating that HNE is able to modulate signaling pathways and protein modification. In the present study, we are interested to test the hypothesis that the accumulation of HNE-protein adducts in normal human chondrocytes will increase under physiological oxidative stress and consequently modulate cell activities. Firstly, we will use proteomics techniques in order to identify specific proteins which will be targeted by HNE binding in normal human chondrocytes under physiological oxidative stress. We will then focus on the enzymatic mechanism involved in HNE-bound proteins accumulation. The glutathione-S-transferase (GST), which detoxifies HNE, acts as a major defense against oxidative stress in cells. We will optimize the chemical and molecular interventions in isolated chondrocytes in order to control HNE-bound proteins levels through GST regulation. Finally, we propose to clarify the significance of increased levels of HNE-bound proteins on cell phenotype and metabolism in human normal chondrocytes.